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SRP072988
Mus musculus
98 Downloadable Samples
Illumina HiSeq 2000
Description
Acinar cells make up the majority of all cells in the pancreas, yet the source of new acinar cells during homeostasis remains unknown. Using multicolor lineage-tracing and organoid-formation assays, we identified the presence of a progenitor-like acinar cell subpopulation. These cells have long-term self-renewal capacity, albeit in a unipotent fashion. We further demonstrate that binuclear acinar cells are terminally differentiated acinar cells. Transcriptome analysis of single acinar cells revealed the existence of a minor population of cells expressing progenitor markers. Interestingly, a gain of the identified markers accompanied by a transient gain of proliferation was observed following chemically induced pancreatitis. Altogether, our study identifies a functionally and molecularly distinct acinar subpopulation and thus transforms our understanding of the acinar cell compartment as a pool of equipotent secretory cells. Overall design: The single-cell RNA-seq library preparation protocol was based on the SMART seq2 protocol (Picelli et al., 2014) with following modifications. Acinar cells were isolated as described in the section Acinar Cell Isolation and Culture and resuspended in DPBS without Ca2+ and Mg2+ (PAN-Biotech). Cells were collected in a volume of 0.5 µL and transferred to a reaction tube containing 4 µL of 6 M guanidine-HCl (Sigma-Aldrich), 0.1% (v/v) Triton X-100 (Sigma-Aldrich) and 1% (v/v) 2-mercaptoethanol (?Sigma-Aldrich). The tube was immediately transferred into liquid nitrogen and kept there for the duration of cell collection. Next, 2.2× RNA SPRI beads (Beckman Coulter) were added directly to the lysis buffer and incubated for 5 min at room temperature. The beads were washed twice with 70% ethanol. Air-dried beads were resuspended in a solution containing 2 µL of H20, 1 µL of oligo(dT) primer, and 1 µL of dNTP Mix (primer and nucleotides used as in Picelli et al., 2014). Twenty-four cells contained ERCC Spike-In RNAs (1:10,000; Mix2, Ambion) Mix in addition to primer and nucleotides. Beads were incubated for 3 min at 72°C, and reverse transcription and PCR (19 cycles) were performed as described by Picelli et al. (2014). PCR product was cleaned up using 0.8× DNA SPRI beads (Beckman Coulter), and air-dried beads were resuspended in 15 µL of H2O. The quality of cDNA library was assessed for each cell on a high-sensitivity DNA Bioanalyzer chip. Subsequent steps (tagmentation, amplification, multiplexing) were done as previously described (Llorens-Bobadilla et al., 2015). The DKFZ Genomics and Proteomics Core Facility conducted sequencing on an Illumina HiSeq2000 sequencer (paired-end 100 bp).
Publication Title
Single-Cell Analysis Uncovers Clonal Acinar Cell Heterogeneity in the Adult Pancreas.
Sample Metadata Fields
Sex, Specimen part, Cell line, Subject
View Samples- Mus musculus
- 6 Downloadable Samples
- Illumina HiSeq 2500
Description
we used technique that allows the molecular characterization of particular neuronal subpopulations based on their neuroanatomical projections and the locations of their cell bodies. This 'retro-TRAP' (translating ribosome affinity purification from retrogradely labeled neurons) approach relies on viral injection into an anatomical area targeted by the neurons of interest, followed by selective precipitation of ribosomes from retrogradely labeled cell bodies, and subsequent RNAseq analysis. Overall design: By comparing the mRNAs enriched in the NGC neurons which are retrogradely labeled due to viral injection into central thalamus, to gene expression of non-labeled surrounding cells in NGC, and then performing a comprehensive bioinformatics analysis of these results, we were able to identify genes enriched in these cells. This procedure allowed us to highlight genes and pathways unique to these neurons with projections ascending to thalamus, as compared to other cells in reticular NucleusGigantocellularis.
Publication Title
Molecular profiling of reticular gigantocellularis neurons indicates that eNOS modulates environmentally dependent levels of arousal.
Sample Metadata Fields
Sex, Specimen part, Cell line, Subject
View Samples- Mus musculus
- 6 Downloadable Samples
- Illumina Genome Analyzer II
Description
We report the application of single-molecule-based sequencing technology for high-throughput profiling of NSC transcriptome. Overall design: Wild type and Sox2-deleted NSC were sequenced; three independent samples from wild type, and three from Sox2-deleted brains (different individual mice).
Publication Title
Mapping the Global Chromatin Connectivity Network for Sox2 Function in Neural Stem Cell Maintenance.
Sample Metadata Fields
Specimen part, Subject
View Samples- Xenopus laevis
- 8 Downloadable Samples
Affymetrix Xenopus laevis Genome Array (xenopuslaevis)
Description
Foregut organogenesis is regulated by inductive interactions between the endoderm and the adjacent mesoderm. We identified genes induced in the foregut progenitors by the adjacent mesoderm.
Publication Title
Sizzled-tolloid interactions maintain foregut progenitors by regulating fibronectin-dependent BMP signaling.
Sample Metadata Fields
Specimen part
View Samples- Homo sapiens
- 35 Downloadable Samples
- IlluminaHiSeq2000
Description
Background: Personalized medicine is predicated on the notion that individual biochemical and genomic profiles are relatively constant in times of good health and to some extent predictive of disease or therapeutic response. We report a pilot study quantifying gene expression and methylation profile consistency over time, addressing the reasons for individual uniqueness, and its relation to N=1 phenotypes. Methods: Whole blood samples from 4 African American women, 4 Caucasian women, and 4 Caucasian men drawn from the Atlanta Center for Health Discovery and Well Being study at three successive 6-month intervals were profiled by RNASeq, miRNASeq, and Illumina Methyl-450 arrays. Standard regression approaches were used to evaluate the proportion of variance for each type of omic measure that is among individuals, and to quantify correlations among measures and with clinical attributes related to wellness. Results: Longitudinal omic profiles are in general highly consistent over time, with an average of 67% of the variance in transcript abundance, 42% of CpG methylation level (but 88% for the most differentiated CpG per gene), and 50% of miRNA abundance among individuals, which are all comparable to 74% of the variance among individuals for 74 clinical traits. One third of the variance can be attributed to differential blood cell type abundance, which is also fairly stable over time, and a lesser amount to eQTL effects, whereas seven conserved axes of covariance that capture diverse aspects of immune function explain over half of the variance. These axes also explain a considerable proportion of individually extreme transcript abundance, namely approximately 100 genes that are significantly up- or down-regulated in each person and are in some cases enriched for relevant gene activities that plausibly associate with clinical attributes. A similar fraction of genes have individually divergent methylation levels, but these do not overlap with the transcripts, and fewer than 20% of genes have significantly correlated methylation and gene expression. Conclusions: People express an “omic personality” consisting of peripheral blood transcriptional and epigenetic profiles that are constant over the course of a year and reflect various types of immune activity. Baseline genomic profiles can provide a window into the molecular basis of traits that might be useful for explaining medical conditions or guiding personalized health decisions. Overall design: Whole blood samples from 12 subjects drawn from the Atlanta Center for Health Discovery and Well Being study at three successive 6-month intervals were profiled by RNASeq, miRNASeq, and Illumina Methyl-450 arrays.
Publication Title
Omic personality: implications of stable transcript and methylation profiles for personalized medicine.
Sample Metadata Fields
No sample metadata fields
View Samples- Saccharomyces cerevisiae
- 2 Downloadable Samples
- Affymetrix Yeast Genome S98 Array (ygs98)
Description
The ARV1-encoded protein mediates sterol transport from the endoplasmic reticulum (ER) to the plasma membrane. Yeast ARV1 mutants accumulate multiple lipids in the ER and are sensitive to pharmacological modulators of both sterol and sphingolipid metabolism. Using fluorescent and electron microscopy, we demonstrate sterol accumulation, subcellular membrane expansion, elevated lipid droplet formation and vacuolar fragmentation in ARV1 mutants. Motif-based regression analysis of ARV1 deletion transcription profiles indicates activation of Hac1p, an integral component of the UPR. Accordingly, we show constitutive splicing of HAC1 transcripts, induction of a UPR reporter and elevated expression of UPR targets in ARV1 mutants. IRE1, encoding the unfolded protein sensor in the ER lumen, exhibits a lethal genetic interaction with ARV1, indicating a viability requirement for the UPR in cells lacking ARV1. Surprisingly, ARV1 mutants expressing a variant of Ire1p defective in sensing unfolded proteins are viable. Moreover these strains also exhibit constitutive HAC1 splicing that interacts with DTT-mediated perturbation of protein folding. These data suggest a component of UPR induction in arv1? strains is distinct from protein misfolding. Decreased ARV1 expression in murine macrophages also results in UPR induction, particularly up-regulation of activating transcription factor-4, C/EBP homologous protein (CHOP) and apoptosis. Cholesterol loading or inhibition of cholesterol esterification further elevated CHOP expression in ARV1 knockdown cells. Thus, loss or down-regulation of ARV1 disturbs membrane and lipid homeostasis resulting in a disruption of ER integrity, one consequence of which is induction of the UPR.
Publication Title
Loss of subcellular lipid transport due to ARV1 deficiency disrupts organelle homeostasis and activates the unfolded protein response.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 9 Downloadable Samples
- Illumina HiSeq 2500
Description
Background. More than one million women in fertile age are infected with Trypanosoma cruzi worldwide. Anti-T.cruzi seropositivity in mothers has been associated with adverse pregnancy outcome but there is still a knowledge gap regarding this effect. Our aim was to compare the gene expression profile of term placental environment from T. cruzi seropositive (SP) and seronegative (SN) mothers. Methods. A RNA-Seq was performed in 9 pools of 2 different placental RNA samples each: 3 belonging to placentas from SN and 6 from SP. Each pool consisted of a binomial of a female/male newborn and a vaginal/caesarean delivery. None of the newborns resulted infected. Results. Only 42 genes showed a significant fold change between SP and SN groups. Among the down-regulated genes were KISS1 and CGB5. In the up-regulated genes group were: KIF12, HLA-G, PRG2, TAC3, FN1 and ATXN3L. To identify pathways significantly associated with maternal T. cruzi-infection, a gene-set association analysis was implemented. The placental environment transcriptomic profile of SP consisted of an enrichment in immunological genes sets (inflammatory response and lymphocytic activation were over-expressed) whereas numerous biosynthetic processes were down-regulated. Conclusions. It is worth noting that several differentially expressed genes in SP placentas code for proteins associated to preeclampsia and miscarriage. This first transcriptomics study in human term placental environment from non-infected deliveries shows a placental response that may affect the faetus while protecting it from the parasite infection; this host response could be responsible for the low rate of congenital transmission observed in human chronic Chagas disease. Background. More than one million women in fertile age are infected with Trypanosoma cruzi worldwide. Anti-T.cruzi seropositivity in mothers has been associated with adverse pregnancy outcome but there is still a knowledge gap regarding this effect. Our aim was to compare the gene expression profile of term placental environment from T. cruzi seropositive (SP) and seronegative (SN) mothers. Methods. A RNA-Seq was performed in 9 pools of 2 different placental RNA samples each: 3 belonging to placentas from SN and 6 from SP. Each pool consisted of a binomial of a female/male newborn and a vaginal/caesarean delivery. None of the newborns resulted infected. Results. Only 42 genes showed a significant fold change between SP and SN groups. Among the down-regulated genes were KISS1 and CGB5. In the up-regulated genes group were: KIF12, HLA-G, PRG2, TAC3, FN1 and ATXN3L. To identify pathways significantly associated with maternal T. cruzi-infection, a gene-set association analysis was implemented. The placental environment transcriptomic profile of SP consisted of an enrichment in immunological genes sets (inflammatory response and lymphocytic activation were over-expressed) whereas numerous biosynthetic processes were down-regulated. Conclusions. It is worth noting that several differentially expressed genes in SP placentas code for proteins associated to preeclampsia and miscarriage. This first transcriptomics study in human term placental environment from non-infected deliveries shows a placental response that may affect the faetus while protecting it from the parasite infection; this host response could be responsible for the low rate of congenital transmission observed in human chronic Chagas disease. Overall design: Serodiagnosis of pregnant women was done by means of conventional serological methods and carried out by the respective health centres based on routine assays. In maternal and umbilical cord blood samples T. cruzi presence was tested using multiplex Real Time PCR as previously described [6]. Maternal infection with other pathogens that produce congenital transmission and adverse pregnancy outcome were considered as exclusion criteria, as well as missing data or incorrect sampling. Fresh normal placentas were obtained after labour from vaginal or caesarean deliveries and placed within 24 hours at 4°C. Each placenta was dissected and the middle section [7] at 2 cm distance from the umbilical cord was isolated and placed into RNAlater solution (Applied Biosystems, Foster City, CA). Total RNA was extracted with TRIzol reagent (Invitrogen, Carlsbad, CA) and stored at -80°C until used. Transcriptomic studies. A RNA-Seq experiment was done in 9 pools of 2 different placental RNA samples each: 3 pools (C1, C2 and C3) belonging to placentas from seronegative mothers (SN) and 6 pools (TC4 to TC9) from seropositive mothers (SP). Each pool consisted of a binomial of a female/male newborn and a vaginal/caesarean delivery. The cDNA Libraries were prepared according to Illumina''s TruSeq Stranded Total RNA with Ribo-Zero Gold for Human and a Hiseq 2.500 Illumina platform with 100 bp paired-end reads was used for sequencing
Publication Title
Alterations in Placental Gene Expression of Pregnant Women with Chronic Chagas Disease.
Sample Metadata Fields
Subject
View Samples- Homo sapiens
- 183 Downloadable Samples
- Illumina HumanRef-8 v3.0 expression beadchip
Description
Maternal smoking has a severe negative effect on all stages of pregnancy that in consequence impairs fetal growth and development. Tobacco smoke-related defects are well established at the clinical level; however, little is known about molecular mechanisms underlying these pathological conditions. We thus employed a genomic approach to determine transcriptome alterations induced by maternal smoking in pregnancy. We assayed gene expression profiles in peripheral blood (M) leukocytes and placentas (PL) of pregnant smokers and those without significant exposure, and in cord blood (D) leukocytes of their babies. Comparative analyses defined significant deregulation of 193 genes in M cells, 329 genes in placentas, and 49 genes in D cells of smokers. These genes were mainly involved in xenobiotic metabolism, oxidative stress, inflammation, immunity, hematopoiesis, trophoblast differentiation, and vascularization. Functional annotation of the deregulated genes outlined processes and pathways affected by tobacco smoke. In smoker newborns, we identified several deregulated pathways associated with autoimmune diseases. The study demonstrates a limited ability of placenta to modulate toxic effects of maternal tobacco use at the gene expression level.
Publication Title
Transcriptome alterations in maternal and fetal cells induced by tobacco smoke.
Sample Metadata Fields
Age, Specimen part, Subject
View Samples- Homo sapiens
- 104 Downloadable Samples
- Illumina HumanRef-8 v3.0 expression beadchip
Description
Passive smoke intake by pregnant women may have detrimental effects such as spontaneous abortion, lower birth weight, stillbirth, and reduced infant lung function. To extend our knowledge on molecular effects of tobacco smoke exposure in pregnancy, we analyzed transcriptome alterations in passive smokers (PS) and compared them to those in active smokers (AS). Using Illumina Expression Beadchip with 24,526 transcript probes, gene expression patterns were assayed in placentas from PS (N=25) exposed to environmental tobacco smoke (ETS) throughout pregnancy and non-exposed (NS) counterparts (N=35), and in cord blood cells from their newborns. The ETS exposure was evaluated by questionnaire disclosure and cotinine measurement in maternal and cord bloods. A total of 196 genes were significantly deregulated in placentas of PS compared to NS. These genes were primary associated with extracellular matrix, apoptosis, blood clotting, response to stress, embryonic morphogenesis, and lipid metabolism. Cord blood of newborns of PS displayed differential expression of 116 genes encoding mainly neuronal factors, regulators of immunologic response, and protooncogenes. Gene ontology analyses highlighted some important biological processes that might be associated with placental insufficiency and fetal growth restriction in PS, such as fatty acid catabolism, coagulation, regulation of growth, and response to steroid hormone stimulus. The study demonstrates that even low dose exposure to ETS during pregnancy leads to the significant deregulation of transcriptional regulation in placental and fetal cells. The data suggest the effect of ETS on the fetus is primary indirect, mediated via deregulation of placental functions. Comparison of PS and AS indicated that ETS exposure and active smoking in pregnancy partly employ the same molecular mechanisms.
Publication Title
Deregulation of gene expression induced by environmental tobacco smoke exposure in pregnancy.
Sample Metadata Fields
Age
View Samples- Homo sapiens
- 14 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
Human diffuse intrinsic pontine gliomas (DIPG) are an aggressive form of pediatric brain tumors that arise in the pons in young children thus resulting in significant morbidity and very poor survival. Recent data suggest that mutations in the histone H3.3 variant are often found in these tumors, though the mechanism of their contribution to oncogenesis remains to be elucidated. Here we report that the combination of constitutive PDGFRA activation and p53 suppression as well as expression of the K27M mutant form of the histone H3.3 variant leads to neoplastic transformation of hPSC-derived neural precursors. Our study demonstrates that human ES cells represent an excellent platform for the modeling of human tumors in vitro and in vivo, which could potentially lead to the elucidation of the molecular mechanisms underlying neoplastic transformation and the identification of novel therapeutic targets.
Publication Title
Use of human embryonic stem cells to model pediatric gliomas with H3.3K27M histone mutation.
Sample Metadata Fields
Specimen part
View Samples